Adsorbents for high mobility group proteins and column for purifying body fluid

ABSTRACT

An adsorbent of high-mobility-group proteins (HMG protein) which can remove HMG protein in body fluid is disclosed. The adsorbent according to the present invention comprises a water-insoluble carrier on which (a) substance(s) having (a) hydrogen-bondable functional group(s) and/or (a) hydrophobic functional group(s) is(are) immobilized.

TECHNICAL FIELD

[0001] The present invention relates to an adsorbent for adsorbinghigh-mobility-group proteins (hereinafter also referred to as “HMGproteins” for short). The present invention is suitably used forimproving the state of sepsis by removing HMG proteins in human blood.

BACKGROUND ART

[0002] HMG proteins are a group of non-histone DNA-binding proteins, andHMG-1, HMG2, HMG14, HMG17, HMG I(Y) and the like are known. HMG havebeen thought to participate in acceleration of transcription and growthof cells by binding to DNA in the cells. It was proved that amphoterindiscovered as a factor which exists on the surface of nerve cells andextends dendrite is HMG-1 that is one of HMG proteins, so that it issuggested that HMG proteins exhibit a wide variety of actions.

[0003] Recently, it was reported that HMG-1 is secreted to the outsideof the cells and acts as a strong mediator of systemic inflammationreaction and septic shock (Wang et al., (1999), Science vol.285, p248).That is, if lipopolysaccharide (LPS) is administered to a mouse, HMG-1level in the serum is drastically increased at 8 to 24 hours afteradministration and the mouse dies. In case of administering purifiedHMG-1 together with LPS, these synergistically act and show lethalactivity, and the lethal action by LPS is inhibited by administering ananti-HMG-1 antibody, so that it was shown that HMG-1 is an importantmediator of endotoxin shock. In patients suffering from sepsis, HMG-1level is drastically increased, especially where the patient dies. HMG-1level in blood is also increased in hemorrhagic shock (Ombrellino etal., (1999), Lancet vol.354, p1446). Further, it has been reported thatproduction of HMG-I(Y) belonging to HMG-1 is induced by stimulation withLPS.

[0004] It was observed that in autoimmune hepatitis, inflammatory boweldisease and in systemic rheumatic diseases, autoantibodies to HMGproteins such as HMG-1, HMG-2, HMG-14 and HMG-17 are produced, so thatit is suggested that HMG proteins participate in these inflammatorydiseases (Sobajima et al., (1997), Clin. Exp. Immunol. vol. 107, p135).Further, it has been reported that HMG proteins participate in growth ofcancers (Taguchi et al., (2000), Nature vol.405, p354).

[0005] Thus, although HMG proteins have functions which are necessary tothe body, in diseases such as sepsis, they are secreted in excess to theoutside of the cells to make the disease worse and to make the body die.A candidate of the method for improving the state of the diseases causedby HMG proteins is to administer a drug such as an antibody, which bindsto HMG and inhibits its action. However, in view of the fact that HMGproteins have functions which are necessary in the cells and on thesurfaces of the cells, it is concerned that administration of a druginhibiting HMG activity may cause a serious side effects. Thus, it isdesired to provide means for selectively removing extracellular HMGproteins which are undesirable to the body.

[0006] It has been reported that some substances such as heparin andRAGE, in addition to the above-mentioned antibodies, bind to HMGproteins (Hori et al., (1995), J. Biol. Chem. vol. 270, p25752, and soon). However, it has not hitherto been proposed to use these HMGprotein-binding substances for removing HMG proteins from the body, andno materials which can remove HMG proteins from body fluids are known.

DISCLOSURE OF THE INVENTION

[0007] Accordingly, an object of the present invention is to provide anadsorbent of HMG proteins which can effectively remove HMG proteins inbody fluid.

[0008] The present inventors intensively studied to discover that byimmobilizing a substance having a particular functional group, the HMGproteins in body fluid can be effectively removed, thereby completingthe present invention.

[0009] That is, the present invention provides an adsorbent ofhigh-mobility-group proteins comprising a water-insoluble carrier onwhich (a) substance(s) having (a) hydrogen-bondable functional group(s)and/or (a) hydrophobic functional group(s) is(are) immobilized. Thepresent invention also provides a body fluid-purification column forremoving high-mobility-group proteins, comprising a column, and theadsorbent according to the present invention. The present inventionfurther provides a method for adsorbing high-mobility-group proteins inbody fluid, comprising contacting the adsorbent according to the presentinvention with body fluid so as to adsorb the high-mobility-groupproteins in the body fluid to the adsorbent. The present inventionfurther provides a use of the adsorbent according to the presentinvention for production of adsorbent material for adsorbinghigh-mobility-group proteins in body fluid.

[0010] By contacting the adsorbent of the present invention with thebody fluid, the HMG proteins in the body fluid are effectively adsorbedby the adsorbent so as to eliminate HMG proteins from the body fluid, sothat the extracellular HMG proteins which are undesirable to the bodymay be removed.

BEST MODE FOR CARRYING OUT THE INVENTION

[0011] As described above, the adsorbent of HMG proteins according tothe present invention comprises an adsorbent of high-mobility-groupproteins comprising a water-insoluble carrier on which (a) substance(s)having (a) hydrogen-bondable functional group(s) and/or (a) hydrophobicfunctional group(s) is(are) immobilized.

[0012] Examples of the hydrogen-bondable functional group includecationic functional groups such as primary amino group, secondary aminogroup, tertiary amino group, imino group and quaternary ammonium group;anionic functional groups such as carboxyl group, sulfate groups,sulfonic acid group and phosphate group; and functional groups andstructures having large polarity such as hydroxyl group, thiol group,aldehyde group, carbonyl group, urea bond and thiourea bond.

[0013] Among these, cationic functional groups and anionic functionalgroups are preferred.

[0014] Among the cationic functional groups, amino groups and quaternaryammonium group are especially preferred. Examples of the especiallypreferred amino groups and quaternary ammonium group include thoserepresented by the following Formula (I):

[0015] (wherein R¹, R² and R³ independently represent hydrogen atom or aC₁-C₅ alkyl group, respectively)

[0016] It should be noted that in the present specification and in theclaims, the term “alkyl group” includes both linear and branched alkylgroups.

[0017] Among the groups represented by Formula (I), those wherein R¹, R²and R³ independently represent hydrogen atom or a C₁-C₂ alkyl group,respectively, are more preferred, and those wherein all of R¹, R² and R³represent methyl group are still more preferred. The functional groupsmay be employed individually or in combination.

[0018] As the anionic functional groups, among those described above,carboxyl group, sulfate groups and sulfonic acid group are especiallypreferred. These functional groups may be employed individually or incombination.

[0019] As the hydrophobic functional groups, alkyl groups having notless than 6 carbon atoms, and aromatic groups are preferred. Althoughthere is no specific upper limit of the number of carbon atoms in thealkyl group, it is usually about 30. Examples of the aromatic groupsinclude phenyl group, naphthyl group, and arylalkyl group and aralkylgroup in which phenyl group or naphthyl group is substituted with one ormore alkyl groups (preferably the number of carbon atoms in each alkylgroup is 1 to 30). These functional groups may be employed individuallyor in combination.

[0020] The substance which is to be immobilized on the water-insolublecarrier is not restricted as long as the substance has one or more ofthe above-mentioned functional groups. Examples of the substance includelow molecular synthetic compounds; synthetic polymers; naturallyoccurring low molecular compounds such as amino acids and sugars;saccharides such as oligo saccharides and polysaccharides, andderivatives thereof; peptides, proteins and modified products thereof;nucleic acids such as DNAs and RNAs, and derivatives thereof; and otherphysiologically active substances, biopolymers and compounds originatedfrom microorganisms. In view of the stability to sterilizationtreatment, those having molecular weights of not more than 20,000 arepreferred, and those having molecular weights of not more than 5000 aremore preferred. Especially, those having molecular weights of about 50to 500 are preferred.

[0021] Preferred examples of the above-mentioned substances havingcationic functional groups include amino acids such as lysine andarginine, and peptides consisting of, or containing a large amount of(preferably not less than 30 mol %) these amino acids, for example,polylysine.

[0022] Examples of the substances having anionic functional groupsinclude heparin and dextran sulfate which are sulfated polysaccharidescontaining a number of sulfate groups, as well as derivatives thereof.Since what is necessary is the sulfate group, the term “derivative”includes any derivatives which retain the sulfate groups of the heparinor dextran sulfate. Examples of the preferred substances having carboxylgroup or sulfonic acid group include amino acids such as aspartic acidand glutamic acid, and peptides consisting of, or containing a largeamount of (preferably not less than 30 mol %) these amino acids.

[0023] Preferred examples of the substances having hydrophobicfunctional groups include hydrophobic amino acids such as phenylalanineand tryptophan, and peptides consisting of, or containing a large amountof (preferably not less than 30 mol %) these amino acids.

[0024] Although the density of the above-mentioned functional groups isnot restricted, the number of the above-mentioned functional groups (incases where a plurality of kinds of functional groups are contained, thetotal thereof per 1 g dry weight of the water-insoluble carrier on whichthe functional groups are immobilized is preferably about 1 μmol to 1mmol, more preferably about 10 μmol to 1 mmol.

[0025] A material comprising a water-insoluble carrier and (an)antibody(ies) to HMG protein may also be used as the adsorbent of theHMG protein. Examples of the materials which may suitably be used as thewater-insoluble carrier used in the present invention include syntheticpolymers such as polyamides, polyimides, poly(aromatic vinyl compounds),polyesters, polymethylmethacrylates, polysulfones, polyethylenes,polyvinylalcohols and polytetrafluoroethylenes; and naturally occurringpolymers such as cellulose, collagen, chitin, chitosan and dextran, aswell as derivatives thereof. In addition, inorganic materials such asmetals, ceramics and glass, of which surfaces are covered with anappropriate polymer, and of which surfaces are directly modified mayalso be suitably employed.

[0026] The material used in the present invention may be in the form offibers, hollow fibers, beads, flat membrane, or powder. Those in theform of fibers, hollow fibers or beads, which are also suited forextracorporeal circulation of whole blood in which blood cells andplasma pass through a column without being separated, are preferred. Toincrease the rate of adsorption, porous materials having largecontacting area are preferred. As the beads, since those giving smallpressure loss when being packed in a column, and since those havinglarge surface area are preferred, those having particle sizes of 50 to1000 μm, especially 200 to 700 μm, are preferred.

[0027] The adsorbent according to the present invention may preferablybe one which selectively adsorb HMG protein such that, when theabove-mentioned adsorbent of the present invention is subjected toadsorption treatment of HGM-containing serum, the rate of adsorption ofthe HMG protein is not less than 50%, preferably not less than 80%,still more preferably not less than 90%, still more preferably not lessthan 95%, and the rate of adsorption of albumin in serum is not morethan 20%, preferably not more than 15%, and still more preferably notmore than 10%. Such an adsorbent which selectively adsorb HMG proteinmay be obtained by immobilizing the above-mentioned preferred functionalgroups on the water-insoluble carrier at the above-mentioned preferreddensity, and a plurality of concrete examples are described in theExamples below.

[0028] The term “rate of adsorption” of HMG protein or albumin meanswhat percent of bovine HMG-1 protein in the sample solution is adsorbed,or what percent of serum albumin in the sample solution is adsorbed,when 50 μl of the adsorbent is added to 0.4 ml of sample solutionprepared by adding bovine HMG-1 protein to normal human serum to aconcentration of 600 ng/ml, and then shaking the resulting mixture at37° C. for 2 hours.

[0029] Immobilization of the substance having the above-mentionedfunctional groups on the water-insoluble carrier via covalent bond maybe accomplished by known methods such as reaction with the carrieractivated with cyanogen bromide, polycondensation by carbodiimide,crosslinkage by a divalent reagent which reacts with amino group orthiol group, and crosslinkage by glutaraldehyde. As is apparent from theabove description, those wherein the substance containing theabove-mentioned functional group(s) is bound to the water-insolublecarrier via covalent bond is also included in “water-insoluble carrieron which the substance is immobilized”. Further, those wherein theabove-mentioned functional groups are introduced or included from thestep of synthesizing the water-insoluble carrier is also included in“water-insoluble carrier on which the substance is immobilized”.

[0030] By packing the adsorbent according to the present invention in acolumn, a body fluid-purification column for removinghigh-mobility-group proteins is obtained. In cases where the adsorbentis in the form of fibers, the adsorbent may be packed into the column bymaking the fibers into woven fabric, knit or non-woven fabric, and bystacking and packing the resulting fabric into the column, or by windingthe fabric around a hollow core pipe having pores, and passing thesolution from the inside to the outside.

[0031] By packing the adsorbent according to the present invention intoa column and by making a body fluid containing HMG proteins such asblood or plasma of a patient pass through the column by extracorporealcirculation, therapy of diseases such as sepsis may be carried out. Byusing the column according to the present invention together with a bodyfluid-purification column for adsorbing bacterial components, especiallyhigh therapeutic effect against sepsis is expected to be obtained.Further, the adsorbent of the present invention may also be suitablyused for the therapy of diseases such as cancers and autoimmunediseases.

EXAMPLES

[0032] The present invention will now be described by way of examplesthereof.

[0033] (Measuring Method)

[0034] (1) Rate of Adsorption of HMG-1

[0035] The HMG-1 concentrations of a sample before and after thereaction in each Example were determined by ELISA and the rate ofadsorption was calculated in Example 4 according to the followingequation:

Rate of Adsorption (%)={1-concentration after adsorption(ng/ml)/concentration before adsorption (ng/ml)}×100

[0036] In Examples 1-3, considering the increase in the volume of thesolution by adding an adsorbent with high water content, the rate ofadsorption was approximately calculated according to the followingequation:

Rate of Adsorption (%)=[1-{concentration after adsorption (ng/ml)×0.45(ml)}/{concentration before adsorption (ng/ml)×0.4 (ml)}]×100

[0037] (Note: 0.45=volume (ml) of reaction solution after adsorption,0.4=volume (ml) of reaction solution before adsorption)

[0038] (2) Rate of Adsorption of Albumin

[0039] Serum albumin levels before and after the reaction were measuredby using an automatic blood biochemistry analyzing apparatus (FujiDrychem 5500 commercially available from Fuji Photo Film Co., Ltd), andthe rate of adsorption of albumin was calculated in the same manner asin the calculation of rate of adsorption of HMG-1.

Example 1 Adsorption of HMG-1 by Beads Having Various Kinds ofFunctional Group

[0040] To normal human serum, bovine HMG-1 protein was added to aconcentration of 600 ng/ml. To 0.4 ml of the obtained solution, 50 μl ofcrosslinked agarose beads containing one of the following types offunctional groups were added, and each of the resulting solution wasshaken at 37° C. for 2 hours.

Adsorbent: (1)—N⁺(CH₃)₃, (2)—N³⁰(C₂H₅)₂H, (3)—SO₃—, (4)—COO—,(5)—(CH₂)₇—CH₃, (6)—C₆H₅

[0041] As a control, the solution to which the beads were not added wasshaken for 2 hours in the same way. The results are shown in Table 1.

[0042] Among the water-insoluble carriers having these functionalgroups, those having cationic functional groups (1) and (2), and thosehaving anionic functional groups (3) and (4) exhibited adsorption ofHMG-1 with high efficiency. The hydrophobic functional groups (5) and(6) also showed adsorption. On the other hand, adsorption of albumin bythese adsorbent was small. TABLE 1 Rate of Adsorption Rate of AdsorptionAdsorbent of HMG-1 (%) of Albumin (%) Control 0 0 (1) —N⁺(CH₃)₃ 99 7 (2)—N⁺(C₂H₅)₂H 97 17 (3) —SO₃ ⁻ 97 10 (4) —COO⁻ 88 7 (5) —(CH₂)₇—CH₃ 51 15(6) —C₆H₅ 69 17

Example 2 Adsorption by Physiologically Active Substance-ImmobilizedBeads

[0043] Using the crosslinked beads on which one of the variousphysiologically active substances shown in Table 2 was immobilized, theadsorption experiments of HMG-1 in normal human serum were carried outin the same manner as in Example 1. As shown in Table 2, the HMG-1 inthe serum was adsorbed and removed with high efficiency by (2) heparin,(4) polyL-lysine, (5) dextran sulfate or (7) histamine, and adsorptionof albumin which is necessary to the body was small.

[0044] Each adsorbent was prepared as follows: One gram of crosslinkedagarose beads activated with cyanogen bromide were mixed with 10 ml ofeach physiologically active substance solution with a concentration of 5mg/ml in 0.1 M aqueous sodium hydrogen carbonate solution containing 0.5M sodium chloride, and the resulting mixture was allowed to react at 4°C. for 24 hours. Then ethanolamine was added to 0.1 M and the resultingmixture was allowed to react at room temperature for 2 hours, therebyblocking the non-reacted active groups. The obtained beads were washedwith distilled water and used in the experiments. TABLE 2 Rate ofAdsorption Rate of Adsorption Adsorbent of HMG-1 (%) of Albumin (%)Control 0 0 (1) DNA-agarose 39 13 (2) Heparin-agarose 98 17 (3)Polymixin B-agarose 30 17 (4) Poly-L-lysine-agarose 97 17 (5) Dextransulfate 91 13 (6) Histamine-agarose 72 10

Example 3 Adsorption by Amino Acid-Immobilized Beads

[0045] Using the crosslinked beads on which one of the variousphysiologically active substances shown in Table 2 was immobilized, theadsorption experiments of HMG-1 in normal human serum were carried outin the same manner as in Example 1. As shown in Table 3, (2)arginine-immobilized material showed high adsorption, and (1)lysine-immobilized material and (2) tryptophan-immobilized material alsoshowed adsorption. Adsorption of albumin by these materials was small.

[0046] Each adsorbent was prepared as follows: One gram of crosslinkedagarose beads activated with cyanogen bromide were mixed with 10 ml ofeach amino acid solution with a concentration of 5 mg/ml in 0.1 Maqueous sodium hydrogen carbonate solution containing 0.5 M sodiumchloride, and the resulting mixture was allowed to react at 4° C. for 24hours. Then ethanolamine was added to 0.1 M and the resulting mixturewas allowed to react at room temperature for 2 hours, thereby blockingthe non-reacted active groups. The obtained beads were washed withdistilled water and used in the experiments. TABLE 3 Rate of AdsorptionRate of Adsorption Adsorbent of HMG-1 (%) of Albumin (%) Control 0 0 (1)L-lysine-agarose 52 10 (2) L-arginine-agarose 83 13 (3)L-phenylalanine-agarose 31 13 (4) L-tryptophan-agarose 57 10

Example 4 Adsorption by Heparin-immobilized Fibrous Carrier

[0047] Sea-island type composite fibers (thickness: 2.6 denier, numberof islands: 16) described in U.S. Pat. No. 4,661,260 consisting of 50parts by weight of sea component (a mixture of 46 parts by weight ofpolystyrene and 4 parts by weight of polypropylene) and 50 parts byweight of island component (polypropylene) were mixed with 50 g ofN-methyrol-α-chloroacetamide, 400 g of nitrobenzene, 400 g of 98%sulfuric acid and 0.85 g of paraformaldehyde, and the resulting mixturewas allowed to react at 20° C. for 1 hour. The fibers were then washedwith nitrobenzene and placed in water to stop the reaction. Thereafter,the fibers were again washed with methanol and warm water to obtainchloroacetamide methylated crosslinked polystyrene fibers (hereinafterreferred to as “AMPSt fibers” for short).

[0048] To 300 mg of AMPSt fibers, 8 ml of 1% v/v ethylenediaminesolution in 0.1 M sodium hydrogen carbonate solution was added and themixture was allowed to react at 37° C. for 3 hours, thereby introducingamino groups to the fibers. After washing the obtained aminated fibers,6 ml of 6.7 mg/ml of aqueous heparin solution and 2 ml of1-ethyl-3,3-dimethylaminopropylcarbodiimide hydrochloride solution wereadded and the resulting mixture was allowed to react at room temperaturefor 21 hours to obtain heparinated fibers. The obtained fibers will behereinafter referred to as “Adsorbent A”.

[0049] On the other hand, to 300 mg of AMPSt fibers, 8 ml of 1 mg/mlheparin-albumin conjugate preparation (SIGMA) solution in 0.1 M sodiumhydrogen carbonate solution (pH 9.6) was added and the mixture wasallowed to react at 37° C. for 16 hours. Then the fibers were reactedwith Tris-HCl having a final concentration of 0.1 M so as to block thenon-reacted active groups to obtain heparin-albumin-bound fibers. Theobtained fibers will be hereinafter referred to as “Adsorbent B”.

[0050] To normal human serum, bovine HMG-1 protein was added to aconcentration of 600 ng/ml. To 0.4 ml of the obtained solution, 20 mg ofAdsorbent A or Adsorbent B obtained by the method described above wasadded, and each mixture was allowed to react at 37° C. for 2 hours undershaking. The concentrations of HMG-1 and albumin in the serum before andafter the reaction were measured and the results are shown in Table 4.It was shown that HMG-1 in the serum is adsorbed and removed by thefibers on which heparin is immobilized. The amount of albumin adsorbedby these materials was small. TABLE 4 Rate of Adsorption Rate ofAdsorption Adsorbent of HMG-1 (%) of Albumin (%) Control 0 0 (1)Adsorbent A 61 15 (2) Adsorbent B 89 10

1. An adsorbent of high-mobility-group proteins comprising awater-insoluble carrier on which (a) substance(s) having (a)hydrogen-bondable functional group(s) and/or (a) hydrophobic functionalgroup(s) is(are) immobilized.
 2. The adsorbent according to claim 1,wherein said hydrogen-bondable functional group(s) is(are) (a) cationicfunctional group(s).
 3. The adsorbent according to claim 2, wherein saidcationic functional group(s) is(are) primary amino group, secondaryamino group, tertiary amino group, imino group and/or quaternaryammonium group.
 4. The adsorbent according to claim 3, wherein saidcationic functional group(s) is(are) represented by the followingFormula (I):

(wherein R¹, R² and R³ independently represent hydrogen atom or a C₁-C₅alkyl group, respectively)
 5. The adsorbent according to claim 4,wherein in Formula (I), R¹, R² and R³ independently represent hydrogenatom or a C₁-C₂ alkyl group, respectively.
 6. The adsorbent according toclaim 5, wherein all of R¹, R² and R³ in Formula (I) represent methylgroup.
 7. The adsorbent according to claim 1, wherein saidhydrogen-bondable functional group(s) is(are) (an) anionic functionalgroup(s).
 8. The adsorbent according to claim 7, wherein saidhydrogen-bondable functional group(s) is(are) carboxyl group, sulfategroup, sulfonic acid group, and/or phosphate group.
 9. The adsorbentaccording to claim 1, wherein said hydrophobic functional group(s)is(are) (an) alkyl group(s) having not less than 6 carbon atoms, or (an)aromatic group(s).
 10. The adsorbent according to claim 1, wherein saidsubstance(s) having (a) hydrogen-bondable functional group(s) and/or (a)hydrophobic functional group(s) is(are) (a) peptide(s) or (an) aminoacid(s).
 11. The adsorbent according to claim 3, wherein saidsubstance(s) having (a) hydrogen-bondable functional group(s) is(are)(a) peptide(s) or (an) amino acid(s) having (an) amino group(s) inits(their) side chain(s).
 12. The adsorbent according to claim 11,wherein said substance having (a) hydrogen-bondable functional group(s)is(are) polylysine.
 13. The adsorbent according to claim 7, wherein (a)polysaccharide(s) having (an) sulfate group(s) is(are) immobilized. 14.The adsorbent according to claim 13, wherein said polysaccharide isheparin or dextran sulfate, or a derivative thereof.
 15. An adsorbent ofhigh-mobility-group proteins comprising a water-insoluble carrier onwhich (an) antibody(ies) to said high-mobility-group proteins is(are)immobilized.
 16. The adsorbent according to claim 15, wherein saidadsorbent has a rate of adsorption for high-mobility-group proteins ofnot less than 50%, and has a rate of adsorption for serum albumin of notmore than 20%.
 17. The adsorbent according to any one of claims 1 to 16,wherein said water-insoluble carrier is in the form of fibers.
 18. Theadsorbent according to any one of claims 1 to 16, wherein saidwater-insoluble carrier is in the form of beads.
 19. The adsorbentaccording to any one of claims 1 to 18, which is used for therapy ofsepsis.
 20. A body fluid-purification column for removinghigh-mobility-group proteins, comprising a column, and said adsorbentaccording to any one of claims 1 to 19, which adsorbent is packed insaid column.
 21. The body fluid-purification column according to claim20, by which extracorporeal circulation of whole blood can be attained.22. The body fluid-purification column according to claim 20 or 21,which is used for therapy of sepsis.
 23. A method for adsorbinghigh-mobility-group proteins in body fluid, comprising contacting saidadsorbent according to any one of claims 1 to 18 with body fluid so asto adsorb said high-mobility-group proteins in said body fluid to saidadsorbent.
 24. The method according to claim 23, which is carried out byusing said body fluid-purification column according to any one of claims20 to
 22. 25. The method according to claim 23 or 24, wherein said bodyfluid is blood.
 26. The method according to any one of claims 24 to 25,which is carried out for therapy of sepsis.
 27. The method according toclaim 26, which is carried by using said body fluid-purification columnaccording to any one of claims 20 to 22 together with (a) bodyfluid-purification column(s) which adsorb(s) (a) substance(s) originatedfrom bacteria.
 28. Use of said adsorbent according to any one of claims1 to 18 for production of adsorbent material for adsorbinghigh-mobility-group proteins in body fluid.
 29. The use according toclaim 27, wherein said body fluid is blood.
 30. The use according toclaim 27 or 28, wherein said adsorbent material is for therapy ofsepsis.